Enantiomerically enriched 2-alkyl-5-halopent-4-enecarboxylic acids and their esters are valuable intermediates for preparing pharmaceuticals, such as, for instance, for delta-amino-gamma-hydroxy-omega-arylalkanecarboxamides, which have renin-inhibiting properties and can be used as antihypertensive agents in pharmaceutical preparations.
Esterases are generally employed in the resolution of racemates and asymmetrization.
However, only very few esterases suitable for preparing chiral compounds are commercially available.
The use of esterase extracts from pig liver is known on the preparative scale. Pig liver esterase (PLE) was isolated long ago from natural sources, and its activity has also been known for a long time (Simonds, J. P. (1919) Amer. J. Physiol. 48, 141; Bamann, E. et al. (1934) Hoppe-Seyler Z. 229, 15; Falconer J. S. and Taylor, D. B. (1946) Biochem. J. 40, 831-834).
Various studies have also already been carried out in order to characterize PLE (Heymann, E. and Junge, W. (1979) Eur. J. Biochem. 95, 509-518; Lehner, R. and Verger, T. (1997) Biochemistry 36, 1861-1868).
It has further been possible to show, for example in WO 01/09079, that esterase extracts from pig liver can selectively hydrolyze the (R) enantiomer of methyl 5-chloro-2-(1-methylethyl)-4-pentenoate.
However, the use of such esterase extracts from natural sources, such as pig liver, is associated with disadvantages.
In the first place, the qualities of the different batches vary and thus make it difficult to optimize industrial processes. Secondly, the use of animal resources in the manufacture of pharmaceutical products is undesired because the presence of viruses and prions cannot always be precluded.
For these reasons there is a need to produce recombinant pig liver esterases of standardized quality in microorganisms.
The cloning of putative esterase genes is described for example in FEBS Lett. (1991), 293, 37-41. The first functional expression of an active pig liver esterase enzyme was described for the first time in WO 02/48322.
WO 2004/055177 describes the preparation of further recombinant esterases by site directed mutagenesis of the recombinant pig liver esterase of seq. ID No. 1 (rPLE) from WO 02/48322. As is evident from the description of WO 2004/055177 and from the article authored by the same inventors in Protein Engineering, 16, 1139-1145, 2003, the modifications of the rPLE sequence from WO 02/48322 were chosen so that a recombinant intestinal pig esterase (PICE) disclosed in David et al., (1998) Eur. J. Biochem. 257, 142-148, is obtained.
The resolution of racemic 2-alkyl-5-halopent-4-enecarboxylic esters is not described in any of these articles.